Regulation of CCAAT/enhancer binding protein isoforms by serum and glucocorticoids in the rat intestinal epithelial crypt cell line IEC-6

Abstract

Members of the CCAAT/enhancer binding protein (C/EBP) gene family are expressed in murine intestine. However, little is known about their regulation in intestinal epithelial cells. In an attempt to determine regulatory mechanisms involved in their expression, we examined C/EBP alpha, beta, and delta isoform expression in response to serum and glucocorticoids in the rat intestinal epithelial crypt-derived cell line IEC-6, by Northern blot, transcription run-on assays, indirect immunofluorescence, Western blot, and electrophoretic mobility shift assays. Serum leads to rapid and transient increases in C/EBP alpha and beta mRNA and protein levels by posttranscriptional regulatory mechanisms, without affecting transcriptional initiation. However, C/EBP-specific DNA binding capacity is not affected by serum. Whereas C/EBP alpha expression is not regulated by glucocorticoids, C/EBP beta and delta mRNA and protein levels are rapidly induced. These inductions result from both increased transcription rates and posttranscriptional regulatory mechanisms as well. Moreover, C/EBP beta and delta containing DNA binding complexes are increased by glucocorticoids as determined by supershift assays, in contrast to C/EBP alpha containing complexes. Immunofluorescence studies show cytoplasmic and nuclear localization for C/EBP alpha, in contrast to a restricted nuclear localization for both C/EBP beta and C/EBP delta. These results confirm C/EBP isoforms expression in intestinal epithelial cells. Differential regulation by serum and glucocorticoids as well as different localization of three C/EBP isoforms suggest a role for this class of transcription factors in the control of gene expression in intestinal epithelial cells.